A Mechanism for Nitrogenase Including Loss of a Sulfide

Nitrogenase is the only enzyme in nature that can fix N₂ from the air. The active cofactor of the leading form of this enzyme contains seven irons and one molybdenum connected by sulfide bridges. In several recent experimental studies, it has been suggested that the cofactor is very flexible, and might lose one of its sulfides during catalysis. In this study, the possible loss of a sulfide has been investigated by model calculations. In previous studies, we have shown that there should be four activation steps before catalysis starts, and this study is based on that finding. It was found here that, after the four reductions in the activation steps, a sulfide will become very loosely bound and can be released in a quite exergonic step with a low barrier. The binding of N₂ has no part in that release. In our previous studies, we suggested that the central carbide should be protonated three times after the four activation steps. With the new finding, there will instead be a loss of a sulfide, as the barrier for the loss is much lower than the ones for protonating the carbide. Still, it is suggested here that the carbide will be protonated anyway, but only with one proton, in the E₃ to E₄ step. A very complicated transition state for H₂ formation involving a large structural change was obtained. The combined step, with a loss of H₂ and binding of N₂ , is calculated to be endergonic by +2.3 kcal mol^-1 ; this is in excellent agreement with experiments in which an easily reversible step has been found.